Human T-cell Lymphotropic Virus type I (HTLV-I) was the first human retrovirus to be associated with disease, Adult T-cell Leukemia/Lymphoma (ATL; ref. 1, 2--various references are referred to in parenthesis to more fully describe the state of the art to which this invention pertains. Full bibliographic information for each citation is found at the end of the specification, immediately preceding the claims. The disclosures of these references are hereby incorporated by reference into the present disclosure) and later with HTLV-I Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP; 3). Recently this virus has been associated with arthropathy (ref. 4), uveitis (ref. 5) and infective dermatitis (ref. 6). HTLV-I has been found in almost every region of the world and it is estimated that approximately 10 to 20 million people are infected (ref. 7).
The envelope protein of HTLV-I is composed of an external surface glycoprotein, gp46 and a noncovalently associated transmembrane anchor protein, gp21; both of these are derived from a common precursor, gp63 (ref. 8). The gp46 HTLV-I envelope protein is one of the smallest retroviral envelope proteins known and exhibits little sequence variability (ref. 9, 10, 11). This genetic stability may be a reflection of the limited coding sequence and a need for structural conservation in order to preserve its functionality. While a number of studies have characterized the HTLV-I gp46 protein (refs. 12, 13), it has been difficult to heterologously express recombinant envelope protein in large amounts for use in biochemical and immunological studies. (refs. 40, 41, 42, 43, 44, 45) We have recently described the expression of the entire HTLV-I envelope protein, gp63, in a baculovirus expression system (ref. 14). Although the recombinant protein was expressed in large amounts, it was insoluble and the majority of protein was not completely post-translationally processed. Following successful solubilization of this protein, the soluble and insoluble forms of gp63 have been used to generate human T-cells lines in vitro (ref. 15) and high anti-envelope antibody titres in rabbits (ref. 16). Unfortunately, only non-neutralizing antibodies were induced by the recombinant gp63 protein as either insoluble inclusion bodies (ref. 14) or in its soluble form. This baculovirus-expressed envelope protein thus cannot be in the natural conformation that it is present in the virus and thus is not optimal for vaccine or diagnostic purposes.
Our previous study demonstrated that a recombinant vaccinia virus (RVV E3) containing the HTLV-I coding region for gp46 alone, produced the conformationally correct envelope surface protein, induced neutralizing antibodies in mice (refs. 16, 17) and expressed envelope protein at much higher levels that it did when gp21 was concomitantly expressed in another construct RVV E1 (ref. 17). In this previous work, however, there was no provision of an isolated and purified envelope protein of Human T-cell Lymphotrophic Virus Type I (HTLV-I) devoid of non-envelope proteins of HTLV-I having substantially the same conformation as the envelope protein in native HTLV-I, especially the Tox and p12.sup.I proteins which have demonstrated oncogenic potential.
It would be advantageous to provide a recombinantly-produced, isolated and purified envelope protein of HTLV-I which is devoid of other HTLV-I proteins and having substantially the same conformation as the native protein in high yields and methods of purification of such proteins. Such proteins have use as antigens, immunogenic preparations, including vaccines, as components of diagnostic assays and for the generation of diagnostic reagents. It would also be advantageous to provide human monoclonal antibodies which are HTLV-I envelope protein specific and substantially non-binding to HTLV-I envelope protein in a denatured form.